Melrose J, Ghosh P
Raymond Purves Research Laboratories, University of Sydney, Royal North Shore Hospital of Sydney, St. Leonards, N.S.W., Australia.
Biochem Int. 1987 Jul;15(1):117-25.
Human lumbar disc tissue when extracted with 4M GuHCl and subjected to dissociative CsCl density gradient ultracentrifugation yielded trypsin inhibitor activity in the low bouyant density fractions (rho less than or equal to 1.38 g/ml). Disc proteoglycans sedimented in the high bouyant density fractions (rho greater than or equal to 1.5 g/ml). Sephadex G75F gel filtration of the low bouyant density protein fractions afforded a major low molecular weight (Kav = 0.5) trypsin inhibitor pool which was further purified by trypsin affinity chromatography. This latter step facilitated separation of the trypsin inhibitors from neutral proteinase activity also present. The trypsin inhibitor fraction so isolated was shown to possess potent inhibitory activity against a range of human serine proteinases including leukocyte elastase and cathepsin G, urokinase, kallikrein, plasmin and thrombin. Significantly this serine proteinase inhibitor preparation effectively prevented degradation of proteoglycans by a neutral proteinase also isolated from the human intervertebral disc.
人腰椎间盘组织用4M盐酸胍提取后，经CsCl解离密度梯度超速离心，在低浮力密度组分（ρ≤1.38 g/ml）中产生胰蛋白酶抑制活性。椎间盘蛋白聚糖沉淀在高浮力密度组分（ρ≥1.5 g/ml）中。对低浮力密度蛋白组分进行Sephadex G75F凝胶过滤，得到一个主要的低分子量（Kav = 0.5）胰蛋白酶抑制剂池，通过胰蛋白酶亲和层析进一步纯化。后一步骤有助于将胰蛋白酶抑制剂与同样存在的中性蛋白酶活性分离。如此分离得到的胰蛋白酶抑制剂组分显示对一系列人丝氨酸蛋白酶具有强大的抑制活性，包括白细胞弹性蛋白酶、组织蛋白酶G、尿激酶、激肽释放酶、纤溶酶和凝血酶。值得注意的是，这种丝氨酸蛋白酶抑制剂制剂有效地防止了从人椎间盘中分离出的中性蛋白酶对蛋白聚糖的降解。
